The transport of gases across cell membranes is one of the most fundamental of physiological processes-O2 for oxidative metabolism, CO2 for acid-base balance, and NH3 for waste disposal. CO2 retention and hyperammonemia are key components of diseases that are major public health concerns. The traditional dogma had been that all gases cross all cell membranes by diffusing through membrane lipid. However, some membranes are gas impermeable and require protein 'gas channels' such as the aquaporins AQP1 (abundant in red blood cells) and AQP5 (abundant in airway epithelia) to conduct gases such as CO2 and NH3. Movement of these gases through AQPs results in a disturbance of pH in microdomains around the channels that we can measure using a pH microelectrode. However, the mechanism of gas conduction is poorly understood. Molecular dynamic simulations, measurements of the pH beneath an electrode touching the surface (pHS) of a model spherical cell (Xenopus oocytes), as well as a mathematical model addressing these pHS changes have provided the first insights into CO2 and NH3 movement through channels. However, a fundamental understanding of such movements across cell membranes requires more advanced multi-scale mathematical models (microscopic, mesoscopic, sub-macroscopic and macroscopic) in order to elucidate mechanisms of gas permeation in normal and pathological states. The PIs (Drs. Boron, Somersalo, and Tajkhorshid) propose to combine state-of-the-art molecular dynamic simulations and computational modeling with novel experimental studies to develop a predictive mathematical model for permeation of various gases across diverse cell membranes of different protein composition, based on integration of data from complementary methodologies across a range of spatial and temporal scales. We will run molecular dynamic simulations of NH3 and CO2 passage through wild-type, mutant, chemically modified, and metal-bound aquaporins in Aim 1 to predict single channel permeabilities (microscopic scale) that will inform the modeling in Aim 2 and cell physiology in Aim 3. In Aim 2, we will create new computational models of gas transport through single and multiple aquaporins in a lipid bilayer (mesoscopic scale), beneath the pHS electrode (sub-macroscopic scale) and in the whole cell (macroscopic scale). Finally in Aim 3, informed by Aims 1 and 2, we will validate the simulations and models in oocytes using electrophysiological and optical methods.